Multiple Sclerosis (MS) is a common neurological disorder characterized by immune attack on myelin and mylein forming cells of the central nervous system. Current treatments are directed at reducing the immune-mediated inflammatory damage to myelin. Recent observations from neuroimaging and pathological research demonstrate, however, that axonal pathology may also play a large role in the clinical deficits of MS patients, suggesting that the protection of axons may be a potentially important addition to MS therapies. The long-term objective of this proposal is to test the hypothesis that the protection of axons through the inhibition of calcium-activated proteases, or calpains, will ameliorate the clinical deficits in a mouse model of MS. This hypothesis is based on a large volume of data demonstrating the importance of calpain activation in axonal degeneration, and our published experience with preventing axonal degeneration and clinical disease in another animal model of neurological disease. In Aim 1 we will use the model of experimental autoimmune encephalomyelitis (EAE) to investigate the relationship of calpain activation to axonal degeneration and clinical disease. We will measured calpain activation using calpain activity assays and immunocytochemical and immunoblot methods to identify calpain-specific spectrin-breakdown products in tissues. In Aim 2 we will treat animals with EAE with our novel ketoamide calpain inhibitor AK295 and measure attenuation of disease by clinical and patholgical methods. In Aim 3 we will test the ability of AK295 to cross the blood-brain barrier (BBB), since any effective calpain inhibitor will need to get into the brain in sufficient quantities. We will also test the BBB permeability of 10 other novel calpain inhibitor compounds taken from our library of compounds, and compare them to AK295 for BBB permeability. Finally, we will attempt to increase BBB permeability by chemically modifying AK295 and other promising compounds by adding choline and nucleosides in order to take advantage of the inherent BBB transporter systems. These studies of calpain inhibition in a model of MS represent a novel approach to the treatment of this common and devastating neurological disorder. Combined with immunomodulatory therapy, axonal protection by calpain inhibition may significantly change the course of disease for people with MS. [unreadable] [unreadable]